DESCRIPTION (provided by candidate): Bacterial infections are of substantial global concern due to increasing antibiotic resistance. Staphylococcus aureus is a pathogen of alarm because of the ability of the bacterium to infect nearly every site in the human body. This adaptability combined with increasing antibiotic resistance results in the high levels of morbidity and mortality associated with staphylococcal infections. A powerful strategy utilized by vertebrates to combat bacterial pathogens is the sequestration of essential nutrients, a process known as nutritional immunity. Recent work found that during invasive staphylococcal disease mature abscesses are virtually devoid of manganese (Mn). Further work revealed that the sequestration of Mn during staphylococcal infection is dependent on the neutrophil protein calprotectin (CP) a heterodimer of S100A8 and S100A9. In a murine model of infection, CP-deficient mice have increased staphylococcal burdens, suggesting that S. aureus is Mn starved during infection. This idea is further supported by the observation that during infection Mn-dependent bacterial processes are inhibited. In total, these results suggest that CP causes S. aureus to become Mn starved during infection and that this starvation is critical to host defense. Even though vertebrates create an environment virtually devoid of Mn, S. aureus remains capable of causing significant and devastating disease. These observations led us to hypothesize that S. aureus must somehow adapt to the Mn limitation imposed by the host. S. aureus is known to express dedicated Mn transporters, which may help this pathogen resist the effects of vertebrate metal sequestration. However, the increased staphylococcal virulence in CP-deficient mice suggests that expression of Mn transporters by S. aureus is not sufficient to prevent metal starvation. Although it is known that vertebrates sequester Mn, the kinetics of this process have not been elucidated and the extent of Mn starvation imposed on S. aureus remain unknown. Furthermore, how S. aureus adapts to Mn starvation imposed by CP also remains unclear. This application proposes to determine when during infection the host sequesters Mn and the level of metal starvation that is experienced by S. aureus. Additionally, the proposed experiments will determine the contribution of dedicated Mn import systems to resisting CP and elucidate how S. aureus adapts to Mn starvation. The Specific Aims of this application are as follows: Aim 1. Evaluate the kinetics of Mn sequestration during infection and the extent of Mn deficiency experienced by S. aureus. Aim 2. Determine if Mn import systems contribute to the ability of S. aureus to overcome vertebrate Mn sequestration. Aim 3. Determine how S. aureus adapts to Mn limitation imposed by the host. PUBLIC HEALTH RELEVANCE: Bacterial pathogens, such as Staphylococcus aureus, are of serious concern due to their substantial disease burden and increasing antibiotic resistance. To combat these invaders, vertebrates limit the availability of essential nutrients. These studies wil determine the processes that S. aureus uses to overcome the nutrient limiting environment created by the host, facilitating the development of novel therapeutics to treat bacterial infectio.